STUDIES
`
`AND CREATININE
`IN CREATINE
`METABOLISM.
`
`V. THE
`
`METABOLISM
`
`OF CREATlNE.
`
`BY STANLEY
`
`R. BENEDICT
`
`AND EMIL
`
`OSTERBERG.
`
`(From
`
`the Department
`
`of Chemistry,
`New
`
`York
`
`Cornell
`City.)
`
`University
`
`Medical
`
`College,
`
`(Received
`
`for
`
`publication,
`
`March
`
`17, 1923.)
`
`to the
`In 1906 Folin (1) published a memorable contribution
`subject of creatine metabolism in which, as a result of experiments
`upon human subjects, he concluded that
`the animal organism
`cannot convert creatine into creatinine. The conclusion was also
`reached that while creatininc is a waste product, creatine is a food
`and that
`there is no biological relationship between these com-
`pounds. Klercker
`(2) reached quite similar conclusions inde-
`pendently of Folin’s observations.
`In view of the very recent admirable summary of the literature on
`creatine and creatinine given by Hunter (3), it is quite unnecessary
`for us to present an account of the mass of data which has accu-
`mulated since Folin’s
`first contribution.
`A number of in-
`vestigators have reported
`that
`traces of extra creatinine may
`appear in the urine following creatine administration and that
`with very large doses of creatine (Rose and Dimmitt, 4) appreciable
`quantities of extra creatinine may be eliminated. Such findings,
`together with the very excellent and suggestive work of Myers and
`Fine upon the relationship between muscle creatine and urinary
`creatinine elimination, have led many to question seriously Folin’s
`original conclusions. Yet all the evidence presented on the origin
`of urinary creatinine from creatine during the 16 years which have
`elapsed since Folin’s paper, has not been very convincing. Folin
`himself has never abandoned his original position, and there are
`many who would agree with him that practically no positive
`evidence has been offered that creatine is the normal biological
`source of creatinine. Even those most strongly favoring the view
`229
`
`001
`
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`
`

`
`230
`
`Creatine Metabolism
`
`the
`that
`from creatine would have to admit
`that crcatinine arises
`evidence
`for this view must be selected piecemeal here and there,
`wherever
`it can be found.
`Direct,
`clear-cut
`evidence has been
`wholly
`lacking.
`The
`failure
`to
`find conversion
`of appreciable
`amounts
`of creatine
`into creatinine,
`together with
`the fact
`that
`“creatinuria”
`is a definite entity which
`is usually unrelated
`to
`changes
`in the creatinine
`elimination,
`are facts which have made
`Folin’s
`position
`almost
`unassailable
`for
`those who
`forget
`the
`chemical
`relationship
`of the compounds
`involved.
`The work
`reported
`in the present paper was a result of the con-
`clusions
`reached by Behre and Benedict
`(5) that creatinine
`does
`not exist
`in appreciable amounts,
`if at all, in blood, while creatine
`(or some combination
`of it) can be demonstrated
`in this
`fluid, and
`accumulates
`in large amounts after ablation of the kidney
`func-
`tion.
`Prior
`to these
`findings we had held the position
`taken by
`Folin, but
`the work
`on creatine and creatinine
`in blood forced us
`to a different
`view-point.
`We concluded
`that creatine, or some
`simple derivative
`of it, must be the precursor
`of creatinine,
`and
`therefore
`sought
`for
`loopholes
`in previous
`investigations
`which
`might open a way
`to attack
`the problem,
`so that some direct
`answer might be obtained
`to the fundamental
`questions
`involved.
`A careful consideration
`of all the facts available
`led us to conclude
`that there was one basic defect in the strongest argument advanced
`by Folin and his followers
`against
`the origin of urinary
`creatinine
`from creatine.
`Creatine was administered
`and did not
`increase
`the urinary
`creatinine.
`But
`the creatine administered was never
`accounted
`for
`in any way whatever.
`Here
`lay
`the weakness
`in
`Folin’s argument.
`It is true that,
`reasoning
`from
`the known
`facts
`of metabolism,
`it seemed safe to assume
`that a nitrogenous
`com-
`pound would surely give rise to its end-products
`in the body during
`the period
`covered by Folin’s
`experiments.
`Yet Folin himself
`demonstrated
`that
`creatine was
`practically
`or wholly
`unique
`among nitrogen
`compounds
`in that
`it failed
`to reappear
`in any
`form-even
`undetermined
`nitrogen-during
`the period
`of his
`observation.
`Obviously,
`if metabolized at all,
`the creatine must
`ultimately give rise to something.
`Then so long as it remains wholly
`unaccounted for, there is no justification
`for the conclusion
`that it may
`not give rise to creatinine.
`
`002
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`231
`
`the
`
`led to undertake
`
`we were
`As a result of these considerations
`experiments
`reported
`in the present paper.
`The general plan of the work was to bring dogs into approximate
`nitrogenous
`equilibrium
`upon a constant
`diet, and then
`to ad-
`minister
`a small amount
`of creatine daily until an appreciable
`quant,ity of the administered
`creatine was accounted
`for
`in some
`form in the urine.
`The results obtained are, we believe, of definite
`interest
`in connection with
`the basic questions
`of creatine and
`creatinine metabolism.
`of
`the spring and summer
`The work was
`carried out during
`1922. The experimental
`animals were
`healthy
`female
`dogs.
`Each was kept
`in a metabolism
`cage and fed upon a constant diet
`throughout
`the experiment.
`The animals were catheterized
`and
`weighed once daily before
`feeding.
`Periods were marked
`for the
`feces by administration
`of bone-black.
`Total nitrogen was deter-
`mined by the macro Kjeldahl method, using potassium and copper
`sulfates as catalyzers.
`Crcatinine was determined by the original
`Folin m&hod,
`using bichromate
`as standard.
`With
`very
`few
`exceptions
`the urines were diluted
`to the same volume each day
`and the reaction and calorimetric
`reading were
`thus carried out
`Creatine
`in the urine was con-
`under
`rigidly
`fixed conditions.
`verted
`to creatinine
`and determined
`as such by
`the procedure
`commonly
`employed
`in this
`laboratory
`(6).
`The creatine em-
`ployed was prepared
`in the
`laboratory
`from creatinine
`obtained
`from urine (7) and was
`recrystallized
`until
`free from any detectable
`trace of creatinine.
`of the creatine was carefully con-
`The method
`for administration
`sidered. We
`finally adopted oral administration
`by mixing
`the
`creatine with
`the food as the method
`to be tried
`first.
`It seemed
`probable
`that
`the experimental
`period would be long, and the oral
`administration
`was
`far preferable
`from
`the practical
`standpoint
`to
`any
`injection procedure.
`It
`is time-saving,
`and avoids
`infection
`and chance of conversion
`of creatine
`to creatinine
`during sterili-
`zation of the solutions.
`We soon found
`that we could check up the
`question of conversion
`of creatine
`to creatinine
`in the gastro-
`intestinal
`tract, as well as that of absorption
`of the creatine as
`such.
`As we shall show
`later
`in this paper,
`the oral method of
`administration
`is ideal from every standpoint
`for the experiments
`which we carried out upon dogs.
`
`003
`
`Harvest Trading Group - Ex. 1121
`
`

`
`232
`
`Creatine Metabolism
`
`the weighed
`the animal was placed upon
`In each experiment
`diet, and became accustomed
`to
`the cage and
`routine
`of the
`experiments many days before
`the
`first periods which we have
`reported
`in the present paper.
`I to III.
`in Tables
`The results of the experiment’s are recorded
`Table
`I presents a detailed
`report of our
`first experiment.
`In-
`cluding
`the preliminary
`period of 9 days,
`this experiment
`lasted
`128 days.
`There were
`ten periods of 7 days each, during which
`the
`dog received 0.620 gm. of creatine daily
`(containing
`1 molecule of
`water
`of crystallization).
`This was equivalent
`to 0.470 gm.
`expressed
`as creatinine.
`The
`ten periods
`of creatine adminis-
`tration were
`followed by seven after periods of 7 days each.
`We believe
`that a detailed study of Table
`I will be found of
`interest.
`We may
`summarize
`the main
`facts brought
`out as
`:
`follows
`The dog had been on the diet for 24 days before the preliminary
`period reported.
`The nitrogen balance was + 1.3 gm. for a period
`of 7 days preceding
`the
`first period
`recorded
`in the table.
`The
`preliminary
`period
`reported
`shows a plus nitrogen balance of 0.7
`gm. for 7 days.
`The creatinine output
`is quite constant
`for this
`(and the preceding unreported
`periods) and averages 405 mg. per
`day. During
`the 1st week of the creatine administration
`there is no
`apparent effect on the creatinine elimination,
`except
`that on the
`last day of
`the period an increase of about 20 mg. above
`the
`average
`level is to be noted.
`Such a slight
`increase
`is typical of
`The
`results
`previously
`reported
`after
`creatine
`administration.
`nitrogen balance of
`the
`first creatine period
`is +2.7 gm.
`This
`represents
`a retention
`of nitrogen equal to more
`than
`twice
`that
`contained
`in the creatine.
`None of the creatine given appears as
`such
`in the urine.
`Since the urinary
`nitrogen
`is definitely below
`that of the preliminary
`period we may
`infer
`that
`the creatine
`is
`practically
`wholly
`retained
`in some form during
`this period.
`The
`second period of creatine administration
`develops several points
`of real
`interest.
`The creatinine
`in the urine
`is definitely above
`that of the preceding period, but shows
`its maximal height during
`the 1st day.
`The creatinine
`for this second period averages about
`40 mg. higher per day than
`is found during
`the control periods.
`This increase appears
`to us as still too small to warrant
`concluding
`that
`there
`is conversion
`of creatine
`into creatinine, but
`it begins
`to be very suggest,ive.
`
`004
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`233
`
`Nitrogen,
`
`Creatinine,
`
`The
`
`dog
`
`received
`
`daily
`
`Eliminalion
`
`I.
`
`TABLE
`and Creatine
`Administration.
`Dog
`1.
`Female.
`the
`following
`diet:
`
`Crackermeal...............................................
`Evaporated
`milk.
`. .
`Cascin....................................................
`(6.65
`
`gm.
`
`nitrogen)
`
`.
`
`. .
`
`Daily
`
`output
`
`in
`
`the urine.
`
`after Creutine
`
`am.
`150
`. . . . . 100
`24
`
`Remarks.
`
`B
`2 6.
`yj
`.;
`.$
`. .
`a
`T=i
`8
`E
`___-
`Pm. m!7.
`5.711
`412
`5.571
`40:
`6.000
`41:
`5.600
`40:
`5.934
`40:
`5.700
`39;
`5.460
`40:
`5.350
`39i
`5.360
`401
`-~
`5.631
`
`40:
`
`5.746
`5.124
`5.418
`
`5.600
`5.500
`5.644
`5.600
`
`40:
`39:
`39;
`
`40:
`381
`40!
`42f
`
`k/.
`14.4
`
`14.4
`
`14.3
`
`14.4
`
`14.3
`
`--
`k3.626
`5.518
`
`14.4
`
`m!J.
`
`“Vk.
`
`0 Urine
`Feces
`Hair
`Total
`
`=
`=
`=
`=
`
`50.686
`8.282
`0.115
`
`59.083
`
`Food
`0 Balance
`
`N
`
`59.850
`=
`= +0.767
`
`0
`
`0 Urine
`0 Feces
`0 Hair
`
`0 Total
`Food
`
`N
`
`0 Creatine
`
`Total
`Balance
`
`=
`=
`=
`
`=
`=
`
`38.626
`6.316
`0.992
`
`---
`45.034
`46.55
`
`N =
`
`1.215
`
`---
`47.765
`=
`= S2.731
`
`Preliminary
`days
`creatine
`
`period
`duration.
`given.
`
`9
`of
`No
`
`Average
`
`daily
`
`output.
`
`of
`
`creatine
`0.620
`
`period
`First
`administration.
`gm. of creatine(GHsNs-
`with
`Oz. H20)
`was mixed
`This
`the
`food
`each
`day.
`quantity
`of
`crcatine
`would
`represent
`0.470
`gm.
`of creatininc.
`
`2,81!
`40
`
`2
`1
`-
`
`0
`0
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`005
`
`Harvest Trading Group - Ex. 1121
`
`

`
`234
`
`Creatine Metabolism
`
`TABLE
`
`I--Continued.
`
`Daily
`
`output
`
`in
`
`the
`
`urine
`
`2 P
`2
`2 .eo
`B
`kg.
`14.4
`
`14.5
`
`14.4
`
`14.6
`
`15.0
`
`14.7
`
`14.8
`
`14.9
`
`14.8
`
`-
`
`-
`
`RCXIXWkS.
`
`period
`Second
`administration.
`of
`w.
`given
`daily
`
`of creatine
`0.620
`was
`food.
`
`creatine
`in
`the
`
`Total
`Average
`
`for
`
`7 days.
`for
`1 day.
`
`of continued
`period
`Third
`creatine
`administration.
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`Fourth
`tinued
`istration.
`
`period
`creatine
`
`con-
`of
`admin-
`
`0m.
`5.74(
`5.57:
`5.38t
`5.03(
`5.21t
`5.27(
`5.23!
`
`mu.
`45c
`44:
`45(
`44E
`45(
`44:
`44
`
`nw.
`
`um.
`
`Intake
`
`0
`0
`32 Output
`83
`113 Balance
`130
`147
`
`7.431
`5.341
`
`I, 121
`44f
`
`505
`101
`
`5.12‘
`5.04
`5.20
`5.15
`5.04(
`5.12‘
`5.00(
`
`442 1
`44: 5
`45( 1
`46I 1
`45( 1
`45; i
`46( 1
`
`Intake
`
`139
`169
`164 Output
`163
`203 Balance
`209
`204
`
`5.681
`5.09:
`
`I, 161
`45:
`
`1,251
`178
`
`5.26,
`5.05’
`5.23,
`5.29
`5.04
`5.01’
`5.09
`
`46t
`47;
`47:
`47:
`46(
`48:
`48:
`
`Intake
`
`output
`
`209
`221
`239
`253
`245 Balance
`254
`254
`
`=
`
`=
`
`47.765
`
`44.036
`
`=
`
`f3.729
`
`=
`
`=
`
`47.765
`
`42.049
`
`= $5.716
`
`=
`
`=
`
`47.765
`
`42.252
`
`= $5.513
`
`3
`15.99
`5.14
`
`3, 32
`47‘
`
`1,675
`239
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`006
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`235
`
`TABLE
`
`I-Continued.
`
`Daily
`
`output
`
`in
`
`the urine.
`
`i
`
`Remarks
`
`WJ.
`494
`
`T7.
`242
`
`Intake
`
`gm.
`=
`
`47.765
`
`period
`Fifth
`creatine
`
`of continued
`administration.
`
`kg.
`15.0
`
`15.3
`
`15.1
`
`15.3
`
`15.4
`
`15.3
`
`15.4
`
`15.:
`
`i
`
`15.E
`
`i
`
`gm.
`5.151
`
`5.067
`4.800
`4.700
`4.760
`4.550
`4.760
`
`4.900
`4.927
`4.970
`4.956
`5.166
`5.151
`5.040
`-_
`35.110
`5.015
`
`=
`
`39.875
`
`= +7.890
`
`=
`
`=
`
`47.765
`
`40.054
`
`= +7.711
`
`=
`
`=
`
`47.76:
`
`41.Oli
`
`= +6.74E
`
`Total
`Average
`
`for
`
`7 days.
`for
`1 day.
`
`period
`Sixth
`creatine
`
`of continued
`administration.
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`Seventh
`tinued
`istration.
`
`period
`creatine
`
`con-
`of
`admin-
`
`481
`494
`494
`50(
`491
`5Ot
`
`248
`230 Output
`217
`223 Balance
`204
`198
`
`3 3.788
`
`4.826
`
`3,47(
`49!
`
`1,562
`223
`
`5.040
`5.040
`4.760
`4.731
`4.788
`4.861
`4.760
`
`50t
`52f
`51:
`49z
`5ot
`481
`5Ot
`
`Intake
`
`224
`230
`258 Output
`242
`218 Balance
`229
`204
`
`2 13.980 3, 53: 1,605
`
`4.854
`50‘
`229
`
`51!
`501
`501
`501
`521
`51,
`52
`
`Intake
`
`output
`
`198
`192
`186
`211
`210 Balance
`237
`245
`
`3,59
`51
`
`1,479
`211
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`007
`
`Harvest Trading Group - Ex. 1121
`
`

`
`236
`
`Creatine Metabolism
`
`TABLE
`
`I-continued.
`
`Daily
`
`output
`
`in
`
`the urine.
`
`kg.
`15.8
`
`,
`
`15.0
`
`I
`
`15.5
`
`15.5
`
`15.7
`
`15.7
`
`.-
`
`15.7
`
`15.7
`
`15.6
`
`15.i
`
`om.
`5.012
`5.026
`5.375
`5.823
`5.754
`5.684
`5.544
`
`mo.
`5oi
`50(
`54(
`56C
`55:
`53:
`54c
`
`mo.
`205
`187
`203
`192
`224
`285
`286
`
`8.218
`3
`5.459
`
`,734
`53:
`
`1,582
`226
`
`5.375
`5.081
`5.124
`5.070
`5.320
`5.067
`4.760
`
`5.797
`3,
`5.113
`
`5.012
`4.927
`1
`5.110
`/
`5.067
`I
`5.460
`5.290
`!
`I
`5.194
`
`-
`3t
`5.060
`5.151
`
`52E
`51s
`533
`52E
`52E
`533
`51s
`
`6%
`52E
`
`526
`526
`540
`570
`555
`54a
`533
`
`790
`54:
`
`284
`253
`238
`224
`253
`231
`205
`
`,688
`241
`
`210
`210
`224
`201
`294
`270
`238
`
`,647
`23F
`
`Intake
`
`om.
`=
`
`47.765
`
`output
`
`= 44.296
`
`Balance
`
`= +3.469
`
`Eighth
`tinued
`istration.
`
`period
`creatine
`
`of con-
`admin-
`
`Intake
`
`=
`
`47.765
`
`output
`
`= 41.771
`
`Balance
`
`= $5.994
`
`Intake
`
`output
`
`=
`
`47.765
`
`= 42.310
`
`Balance
`
`= +5.455
`
`for 7 days.
`Total
`Average
`for 1 day.
`
`Ninth period of continued
`creatine administration.
`
`for 7 days.
`Total
`Average
`for 1 day.
`
`Tenth period of continued
`creatine administration.
`
`-
`for 7 days.
`Total
`Average
`for 1 day.
`
`008
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`237
`
`-
`
`Daily
`
`d
`m
`‘6
`a’
`.60
`;
`
`ko.
`15.7
`
`15.6
`
`15.7
`
`15.6
`
`*m.
`5.25C
`5.012
`5.29a
`5.375
`5.32C
`5.110
`5.46C
`-
`6.817
`3
`5.259
`
`5.264
`4.703
`4.941
`4.816
`4.563
`5.466
`5.193
`
`4.940
`3
`4.991
`
`5.04C
`5.‘123
`4.983
`4.900
`5.096
`5.488
`5.18C
`_
`3
`m5.81C
`5.116
`
`15.6
`
`15.7
`
`15.7
`
`15.7
`
`d
`2
`.$
`m
`6
`_-
`m.0.
`519
`513
`570
`555
`540
`506
`506
`--
`,709
`529
`--
`494
`482
`488
`500
`488
`494
`506
`--
`,452
`493
`
`500
`506
`494
`488
`482
`500
`494
`
`,464
`494
`
`TABLE
`
`I-continued.
`
`in
`
`the urine.
`
`Intake
`
`output
`
`0m.
`
`=
`
`46.550
`
`=
`
`42.745
`
`Balance
`
`= +3.805
`
`Intake
`
`output
`
`=
`
`46.550
`
`=
`
`41.370
`
`Balance
`
`= +5.1a
`
`output
`-
`
`a
`I
`8
`&T
`HI
`a!‘; ., 4
`2;
`‘2 I
`zc
`6’
`
`mo
`
`: 36
`0
`0
`0
`0
`0
`0
`-_
`36
`:
`
`-.
`0
`0
`0
`
`--
`
`Intake
`
`0
`
`output
`
`=
`
`=
`
`46.550
`
`42.375
`
`0
`
`Balance
`
`= +4.175
`
`0
`-.
`0
`
`Remarks.
`
`after
`First
`creatine
`
`period.
`given.
`
`No
`
`Total
`Average
`
`for
`
`7 days.
`for
`1 day.
`
`Second
`creatine
`
`period.
`after
`given.
`
`No
`
`Total
`Average
`
`for
`
`7 days.
`for
`1 day.
`
`after
`Third
`creatine
`
`period.
`given.
`
`No
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`009
`
`Harvest Trading Group - Ex. 1121
`
`

`
`238
`
`Creatine Metabolism
`
`TABLE
`
`I-Ccncluded.
`
`Daily
`
`output
`
`in
`
`the urine.
`
`T
`
`Remarks.
`
`Intake
`
`output
`
`Qm.
`=
`
`46.550
`
`=
`
`44.340
`
`Balance
`
`=
`
`f2.210
`
`Fourth
`creatine
`
`period.
`after
`given.
`
`No
`
`a
`3
`3
`g
`3
`3
`
`w
`
`I.
`
`0
`
`0
`
`--
`
`--
`
`ww.
`506
`466
`486
`455
`463
`450
`440
`
`ti -3
`“0
`c’ .$
`s
`kg.
`15.7
`
`15.7
`
`15.7
`
`15.7
`
`gm.
`5.29(
`5.47:
`5.57(
`5.334
`5.41(
`5.32(
`5.221
`
`17.62~
`5.37i
`
`5.10(
`5.04(
`5.20;
`5.32(
`4.9s:
`5.07(
`
`Total
`Average
`
`for
`
`7 days.
`for
`1 day.
`
`after
`Fifth
`creatine
`
`period.
`given.
`
`No
`
`Intake
`
`output
`
`=
`
`=
`
`39.900
`
`37.196
`
`0
`
`Balance
`
`= $2.704
`
`,266
`466
`_-
`441
`455
`468
`463
`440
`455
`~-
`:, 722
`453
`__-
`440
`450
`440
`440
`445
`445
`440
`
`10.72(
`5.12(
`
`5.04(
`5.40(
`5.37:
`5.49(
`5.51;
`5.29
`5.39(
`
`17.50
`5.35
`
`I, 100
`442
`
`5.23!
`5.26‘
`5.12‘
`5.321
`5.24:
`5.29
`5.34;
`
`435
`440
`435
`435
`442
`426
`426
`
`36.82‘
`5.261
`
`1,039
`434
`
`15.8
`
`15.7
`
`15.8
`
`15.8
`
`15.8
`
`15.5
`
`15.8
`
`-.
`
`--
`
`0
`
`-.
`
`-I-
`
`O
`
`Intake
`
`output
`
`=
`
`=
`
`46.550
`
`43.965
`
`Balance
`
`= +2.585
`
`Intake
`
`output
`
`=
`
`=
`
`46.550
`
`43.910
`
`Dalance
`
`= +2.640
`
`Total
`Average
`
`for
`
`6 days.
`for
`1 day.
`
`after
`Sixth
`creatine
`
`period.
`given.
`
`No
`
`Total
`Average
`
`for
`
`7 days.
`for
`1 day.
`
`Seventh
`creatine
`
`period.
`after
`given.
`
`No
`
`Total
`Average
`
`for 7 days.
`for
`1 day.
`
`010
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`239
`
`On the 3rd day of this period, or the 10th day of creatine ad-
`ministration,
`a scarcely detectable quantity
`of creatine
`(32 mg.)’
`appears
`in the urine.
`The creatine
`in the urine rapidly
`increases,
`and within
`5 days has reached 147 mg. for 1 day. Here
`is direct
`evidence that creatine given even in small amounts, and apparently
`completely
`utilized,
`is retained, at least partially,
`as such
`in the
`organism
`for a considerable period.
`Furthermore, many days are
`required
`to fill the creatine
`reservoir,
`indicating
`a very slow
`rate
`of metabolism
`for creatine.
`The nitrogen balance
`for the second
`period of creatine
`feeding
`is +3.7 gm.,
`representing
`a storage of
`nitrogen of more
`than
`three
`times
`the quantity
`contained
`in the
`creatine
`retained.
`This
`result
`begins
`to suggest
`that
`creatine
`may cause nitrogen storage
`in the body
`far beyond
`that contained
`in the crcatine
`itself.
`Further
`data show
`t.hat there can be little
`question of the correctness
`of this view.
`throws
`I)
`(Table
`The
`third period of creatine administration
`output
`further
`light upon
`the questions
`raised.
`The creatinine
`frequent
`rises.
`True,
`it
`rises grudgingly
`and unwillingly,
`with
`minor drops.
`This picture
`is typical of all our experiments
`in this
`connection.
`But
`the period as a whole shows a definite
`increase
`in creatinine excretion.
`The figure
`is now about 50 mg. per day
`(12 per cent) above
`the control period,. and we should almost
`feel
`justified
`in concluding
`definitely
`that
`creatine
`gives
`rise
`to
`creatinine
`in
`the organism,
`but with
`the proviso
`that
`this
`is
`apparently
`not a simple direct conversion.
`Definite settlement of
`this question will come
`later.
`The creatine output
`in the urine
`continues
`to rise, reaching about 200 mg. per day toward
`the end
`of
`the
`third
`period of creatine administration.
`Evidently
`the
`creatinc
`reservoir
`can retain
`less and less of the ingested creatine,
`but more
`than 50 per cent of that given
`is st’ill retained.
`The
`nitrogen balance
`for
`this period
`is + 5.7 gm., and the weight of
`the dog is definitely
`increasing.
`We arc
`inclined
`to believe
`that
`the nitrogen
`retention
`and
`increase
`in weight
`arc due
`to
`the
`creatine administered,
`since during
`the 33 days before creatine
`administration
`this animal did not change in weight a total of more
`than 100 gm., and the fluctuations
`were both sides of 14.4 kilos.
`
`* Throughout
`its equivalent
`
`paper
`this
`in creatinine.
`
`the
`
`creatine
`
`found
`
`in
`
`the urine
`
`is expressed
`
`as
`
`011
`
`Harvest Trading Group - Ex. 1121
`
`

`
`240
`
`Creatine Metabolism
`
`feeding
`finding
`
`the weight of the
`is at
`least
`very
`
`the first 21 days of the creatine
`During
`animal has increased
`600 gm. The
`suggestive.
`the
`,administration
`creatine
`period of
`fourth
`the
`During
`in the urine shows
`the same continued
`increase as for
`creatinine
`The average
`for the period shows a
`the two preceding periods.
`daily elimination
`of 70 mg. of creatinine
`(17 per cent) above
`the
`The urinary
`creatine
`con-
`daily average
`for
`the control periods.
`This
`figure
`tinues
`to rise, and shows a daily average of 239 mg.
`is nearly maximal
`for
`the entire
`ten periods of creatine adminis-
`tration, and is maintained
`approximately
`constant as long as the
`creatine
`feeding
`is continued.
`The creatine
`eliminated
`during
`these periods amounts
`to 50 per cent of that given.
`In spite of the increased
`creatinine and creatine
`in the urine, the
`total urinary
`nitrogen
`continues
`low and the nitrogen balance
`for
`the period
`is +5.5 gm.
`The weight
`of the dog shows a slight
`increase
`(100 gm.) over
`the preceding period.
`is
`feeding
`The picture
`for the fifth and sixth periods of creatine
`similar
`to that of the preceding period.
`Creatinine
`in the urine
`continues
`to increase
`slowly,
`so that
`the sixth period shows
`100
`mg. of creatinine
`per day
`in the urine
`in excess of that
`for
`the
`control
`periods.
`The daily
`output
`of creatine
`is somewhat
`irregular,
`but the average
`for the fifth and sixth periods
`is close to
`the average
`for the fourth period.
`Retention
`of nitrogen
`is over
`1 gm. per day for
`the two periods, and the weight of the animal
`increases by about 400 gm.
`further
`feeding shows a slight
`The seventh period of creatine
`rise in creatinine output, while
`the creatine output
`is slightly
`lower
`than
`for the preceding period.
`Nitrogen
`retention
`for the 7 days
`is 6.7 gm.
`The average weight
`of the animal
`for
`this period
`is 1,100 gm. greater
`than
`at
`the beginning
`of
`the creatine
`administration.
`feeding show continued
`The eighth and ninth periods of creatine
`increase
`in the creatinine
`elimination,
`reaching a daily average of
`121 mg. (29 per cent)
`in excess of the control period.
`The creatine
`output
`is practically
`constant.
`Nitrogen
`retention
`for
`the
`two
`periods
`is 9.3 gm. and the weight
`of the dog reaches 1,300 gm.
`above that of the control period.
`
`012
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`241
`
`shows wider
`The tenth and last period of creatine administration
`fluctuations
`in the creatinine
`elimination
`than do the preceding
`periods.
`The maximal
`figure
`for creatinine elimination
`is reached
`during
`this period,
`the avcragc daily output being 541 mg., which
`is 136 mg. or 33 per cent above the daily output
`(405 mg.) during
`the control period.
`The highest
`figure
`for a single day (570 mg.)
`occurs once during
`this and the following
`period.
`The relatively
`wide variations which
`seem to occur at the height of the curve of
`crcatinine
`elimination
`(Period 10 and the
`first after period)
`are
`not apparent during
`the subsequent periods.
`The creatine output
`for the tenth period
`is similar
`to that
`for the ninth.
`the
`At
`the end of the tenth period of creatine administration
`for
`feeding of creatine was discontinued.
`We watched
`the figures
`the following
`period with exceptional
`interest.
`If
`the increased
`crcatinine, which we had found
`in the urine,
`represented
`creatinine
`formed
`in the
`intestinal
`tract
`through
`bacterial
`or other action,
`then the excess creatinine
`in the urine should vanish as promptly,
`or even more promptly,
`than
`the creatine,
`after
`the
`latter was
`discontinued.
`If, on the contrary,
`the extra creatinine
`in the urine
`represented
`the result of a slow, definite metabolism
`of the pre-
`viously
`administered
`creatine,
`the withdrawal
`of
`the creatine
`should have an effect which
`should show
`itself only slowly over a
`considerable period of time.
`are
`after periods
`and subsequent
`The
`results
`for
`the
`first
`beautiful
`and clear-cut.
`The
`first day after withdrawal
`of the
`creatine
`from
`the food
`the creatine of the urine
`fell to a scarcely
`detectable quantity
`(36 mg.) and at no subsequent
`time during
`the
`seven after periods could we detect any creatine
`in the urine.
`The
`creatinine of the urine, on the contrary,
`appears at first
`to have no
`real relationship
`to the administered
`creatine.
`The independence
`of creatine and creatinine
`is again apparent.
`But
`this
`indepen-
`dence is only apparent;
`it is not real.
`Beginning with
`the first
`after period, and continuing
`through
`the six
`following
`periods,
`there
`is a drop
`in the creatinine
`excretion which
`becomes per-
`fectly plain and definite when we consider
`the figures
`for any 7
`day period.
`The drop
`is just as unwilling,
`slow, and slightly
`irregular
`(from day to day) as was
`the rise.
`But it is just as definite
`and clear-cut
`for a number of days considered
`together.
`For
`the
`fifth after period
`the creatinine
`coefficient
`(28) is the same as for
`
`013
`
`Harvest Trading Group - Ex. 1121
`
`

`
`Creatine Metabolism
`
`to store nitrogen, but
`The dog continues
`period.
`the preliminary
`this falls to 2.2 and 2.7 gm. for the last two periods of observation.
`After discontinuing
`the creatine
`the weight of the animal remains
`practically
`constant
`at an average of 15.75 kilos
`for
`the seven
`after periods as against 15.7 kilos
`for
`the first after period.
`to that
`Tables
`II and III
`record
`two
`further experiments
`similar
`recorded
`in Table
`I.
`In order
`to save space the figures are given
`for 7 day periods, and for the daily average of each period,
`instead
`of in full detail as for the experiment
`recorded
`in Table
`I.
`The
`The experiment with Dog 2 is summarized
`in Table
`II.
`entire experimental
`period covered
`twelve periods of 7 days each.
`These
`included
`two preliminary
`control periods,
`five periods
`of
`creatine administration,
`and five after periods.
`0.5 gm. of crea-
`tine was given daily with
`the
`food during
`the creatine periods,
`equivalent
`to 379 mg. per day expressed
`as creatinine.
`This
`represented
`43 mg. per kilo of body weight, which
`is the same per
`kilo dose as that given to Dog 1. There
`is a slight but detectable
`rise
`in crentinine
`elimination
`for
`the
`first
`creatine period, while
`the maximal
`figure
`for creatinine
`in the urine was
`reached
`for
`the
`third creatine period.
`For
`this period
`the creatinine output per
`day was 387 mg.
`This was 92 mg., or 31 per cent, above
`the
`preliminary
`period
`level of 295 mg. of creatinine
`per day.
`This
`high creatinine elimination was approximately maintained
`through
`the last
`two periods of the creatine administration.
`After with-
`drawal of the creatine
`from
`the food the creatinine shows
`the same
`type of very
`slow
`fall observed
`in the experiment with Dog 1
`(Table
`I).
`in the
`(40 mg.) appeared
`In the experiment with Dog 2 creatine
`urine
`first on the 22nd day of creatine administration,
`or 11 days
`later
`than
`in the case of Dog 1, though both
`received
`the same
`quantity
`of creatine per kilo of body weight.
`The quantity
`of
`creatine
`in the urine
`increased
`rapidly
`just as in the preceding
`experiment,
`and during
`the
`fifth
`(and
`last) period of creatine
`administration
`represented
`about 50 per cent of that given,
`just
`as in the preceding experiment.
`Creatine
`(25 mg.) appeared
`for
`the last time
`in the urine on the 1st day of the first after period.
`Nitrogen
`retention
`and
`increase
`in weight of the animal occurs,
`but these are not so great as in the case of Dog 1. Nevertheless,
`there
`is a considerable
`excess of nitrogen
`(above
`that contained
`
`in
`
`014
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`243
`
`increase
`is a slight
`in this animal, and there
`retained
`the creatine)
`The period of creatine administration
`was only one-
`in weight.
`half as long as in the preceding experiment.
`
`Nitrogen,
`
`Creatinine,
`
`II.
`
`TABLE
`and Creatine
`Administration.
`
`Elimination
`
`after
`
`Creatine
`
`The dog
`
`received
`
`daily
`
`the
`
`2.
`Dog
`following
`
`Female.
`diet:
`
`.
`
`.
`
`. . . .
`
`. . . . . .
`
`l7m.
`150
`. 100
`15
`40
`
`meal...............................................
`Cracker
`milk.
`. .
`.
`.
`.
`.
`Evaporated
`Casein....................................................
`Bone
`ash..................................................
`(5.45
`for 7 day periods
`
`The
`
`figures
`
`are
`
`T
`
`am.
`31.774
`$1.916
`il. 650
`
`ma.
`295
`295
`314
`
`am.
`to.24
`to.49
`i-l.24
`
`il.938
`
`2,507
`
`0
`
`358
`
`t1.29
`
`11.8
`11.8
`11.9
`12.1
`12.2
`12.2
`12.3
`12.3
`
`il.777
`S2.222
`S1.833
`31.398
`30.752
`30.981
`31.743
`32.056
`
`2,725
`2,623
`2,610
`2,545
`2,487
`2,403
`2,234
`2,209
`
`0
`837
`1,025
`25’
`0
`0
`0
`0
`
`387
`374
`374
`363
`355
`343
`319
`315
`
`t1.23
`to.90
`to.84
`to. 62
`Il.33
`Il.14
`to.23
`-0.02
`
`-
`
`* Eliminated
`
`1st day
`
`of period.
`
`nitrogen)
`gm.
`unless
`otherwise
`
`indicated.
`
`Remarks.
`
`preliminary
`First
`preliminary
`Second
`Dog
`received
`0.5
`(C4HgN302.H20)
`food.
`Clreatine
`period.
`“
`“
`‘I
`
`given
`
`as
`
`“
`“
`“
`
`period.
`period.
`gm.
`creatine
`daily with
`the
`
`in preceding
`
`“
`‘I
`“
`
`X0 creatine
`I‘
`“
`“
`“
`“
`“
`“
`“
`
`“I
`
`given.
`“
`“
`“
`‘I
`
`third experiment,
`of our
`is recorded a summary
`III
`In Table
`carried out upon Dog 3. The experiment
`covers a period of 92
`days.
`450 mg. of creat,ine, equivalent
`to 341 mg. expressed
`as
`
`015
`
`Harvest Trading Group - Ex. 1121
`
`

`
`244
`
`Creatine Metabolism
`
`feeding,
`the period of creatine
`creatinine, were given daily during
`which
`lasted 66 days.
`There was an after period of 20 days.
`The
`
`Nitrogen,
`
`Creatinine,
`
`The
`
`dog
`
`received
`
`daily
`
`the
`
`Elimination
`
`III.
`
`TABLE
`and Creatine
`Administration.
`Dog
`3.
`Female.
`following
`diet:
`
`ajter
`
`Creatine
`
`Crackermeal...............................................
`Evaporated
`milk.
`. . . . . .
`Casein....................................................
`(4.45
`1 )eriods
`
`The
`
`figures
`
`are
`
`for 7 day
`
`.
`
`9m.
`90
`. . . . . . . 100
`15
`
`.
`
`.
`
`.
`
`.
`
`.
`
`.
`
`. .
`
`.
`
`.
`
`nitrogen)
`gm.
`unless
`otherwise
`
`indicated.
`
`kg.
`9.1
`9.1
`
`9m.
`22.188
`24.052
`
`m9.
`1,662
`2,039
`
`m*.
`
`0
`27*
`
`m9.
`277
`291
`
`9m.
`f0.300
`f2.092
`
`9.1
`
`25.646
`
`2,2241,092
`
`317
`
`f1.650
`
`9.2
`9.2
`9.1
`9.2
`9.2
`9.2
`9.3
`9.2
`
`24.881
`24.602
`23.739
`23.630
`23.731
`24.121
`23.283
`9.980
`
`1,288
`2,277
`1,189
`2,298
`2,3441,371
`2,301
`1,324
`2,347
`1,270
`2,367
`I, 204
`2,364
`1,150
`1,001
`510
`
`325
`328
`335
`328
`335
`338
`337
`333
`
`+2.361
`+2.723
`f3.667
`f3.689
`f3.520
`+3.145
`f3.574
`+1.584
`
`9.2
`
`19.517
`
`1,984
`
`201
`
`330
`
`t2.745
`
`9.1
`9.0
`
`24.266
`25.474
`
`2,181
`2,058
`
`0
`0
`
`311
`294
`
`f2.058
`j-O.281
`
`* Eliminated
`t Eliminated
`
`day
`last
`1st day
`
`of period.
`of period.
`
`0.450
`
`period.
`gm.
`daily
`
`creatine
`with
`
`as
`
`in preceding
`
`“
`“
`“
`“
`“
`“
`“
`Creatine
`period.
`No
`
`given
`
`creatine
`
`given
`
`preliminary
`6 day
`received
`Dog
`(CaHpNaOz.HsO)
`the
`food.
`Creatine
`period.
`“
`“
`“
`“
`“
`“
`“
`
`“
`“
`“
`“
`“
`“
`“
`
`6
`
`period.
`3 day
`as in preceding
`day
`period.
`given.
`No
`creatine
`“
`“
`
`given.
`“
`
`016
`
`Harvest Trading Group - Ex. 1121
`
`

`
`S. R. Benedict and E. Osterberg
`
`245
`
`to those of the prc-
`are quite similar
`of the experiment
`results
`shows
`a slight
`rise
`ceding experiments.
`The creatinine
`output
`during
`the
`first
`creatine period.
`The rise continues, with
`slight
`fluctuations,
`and the creatinine output
`reaches a maximal
`figure of
`338 mg. per day for
`the eighth period of creatine administration.
`This
`figure
`is 61 mg., or 22 per cent, above
`the control period.
`Creatine
`first appeared
`in the urine on the 7th day of creatine
`feed-
`ing, and increased
`rapidly as in the preceding experiments.
`When
`the creatine output was at its height, about 54 per cent of the
`creatine
`given was excreted
`in the urine as such. When
`the
`creatine
`feeding
`is s